Glycosaminoglycans, such as heparin, heparan sulfate, dermatan sulfate, chondroitin sulfate and hyaluronic acid, are biopolymers industrially extracted from different animal organs.
In particular heparin, principally obtained by extraction from intestinal pig mucosa or bovine lung, is a mixture of chains consisting of repeating disaccharide units formed by an uronic acid (L-iduronic acid or D-glucuronic acid) and by an amino sugar (glucosamine), joined by α-1→4 or β-1→4 bonds. The uronic acid unit may be sulfated in position 2 and the glucosamine unit is N-acetylated or N-sulfated and 6-O sulfated. Moreover, glucosamine contains a sulfate group in position 3 in an amount of about 0.5%. Heparin is a polydisperse copolymer with a molecular weight ranging from about 3,000 to about 30,000 D. In particular, extractive heparin contains about 70% iduronic units and about 30% glucuronic units (Casu B., Adv. Carb. Chem. Biochem., 1985, 43, 51-134).
Besides the main anticoagulant and antithrombotic activities, heparin also exerts antilipemic, antiproliferative, antiviral, anticancer and antimetastatic activities. To satisfy the major request of starting material for these new therapeutic areas a new alternative route of production different from the extractive ones from animal tissues is necessary.
The natural biosynthesis of heparin in mammalians and the properties of this product have been described by Lindahl et al. in Lane D. and Lindahl U. (Eds.) “Heparin-Chemical and Biological Properties; Clinical Applications” Edward Arnold, London, 1986, 159-190 and Lindahl U. et al., TIBS, 1986, 11, 221-225.
The sequence formed by the pentasaccharide region of linkage for Antithrombin III (ATIII) named active pentasaccharide
(J. Choay et al., Ann. N.Y. Acad. Sci., 1981, 376, 644-649), that is the structure needed for the high affinity binding of heparin to ATIII, is fundamental for heparin activity. This sequence contains one glucosamine unit sulfated in position 3, that is not normally present in the other parts of the heparin chain. Beside the activity through ATIII, heparin exerts its anticoagulant and antithrombotic activity through the activation of heparin cofactor II (HCII) and a selective inhibition of thrombin. It is known that the minimum saccharidic sequence necessary for HCII activation is a chain containing at least 24 monosaccharides (Tollefsen D. M. Seminars in Thrombosis and Hemostasis, 1990, 16, 66-70).
Heparin is a leading, widely used medicament for treating vascular disorders such as deep vascular thrombosis, or to prevent thrombosis (see Lassen M. R. et al., “Efficacy and safety of prolonged thromboprophylaxis with a low molecular weight heparin (Dalteparin) after total hip arthroplasty” Thrombosis Research, 1998, 89, 281-287 and, more recently, Motsch J. et al. “Update in the prevention and treatment of deep vein thrombosis and pulmonary embolism” Current Opinion in Anesthesiology, 2006, 19, 52-58), but it is subjected to drawbacks due to its extraction from bovine or porcine tissues, in particular to the risk of the presence of viruses or prions (as set forth in U.S. Pat. No. 5,958,899) or of other toxic glycosaminoglycans which can induce severe, even lethal, effects in patients under heparin anticoagulant treatment (Guerrini M. et al., Nat. Biotechnol., 2008, 26, 669-675). Hence, there is an extreme need of a medicament having the same biologic characteristics as heparin without the potential drawbacks thereof.